Identification of Quantitative Trait Loci Controlling Root Morphological Traits in an Interspecific Soybean Population Using 2D Imagery Data.

Roots are the hidden and most important part of plants. They serve as stabilizers and channels for uptaking water and nutrients and play a crucial role in the growth and development of plants. Here, two-dimensional image data were used to identify quantitative trait loci (QTL) controlling root traits in an interspecific mapping population derived from a cross between wild soybean 'PI366121' and cultivar 'Williams 82'. A total of 2830 single-nucleotide polymorphisms were used for genotyping, constructing genetic linkage maps, and analyzing QTLs. Forty-two QTLs were identified on twelve chromosomes, twelve of which were identified as major QTLs, with a phenotypic variation range of 36.12% to 39.11% and a logarithm of odds value range of 12.01 to 17.35. Two significant QTL regions for the average diameter, root volume, and link average diameter root traits were detected on chromosomes 3 and 13, and both wild and cultivated soybeans contributed positive alleles. Six candidate genes, Glyma.03G027500 (transketolase/glycoaldehyde transferase), Glyma.03G014500 (dehydrogenases), Glyma.13G341500 (leucine-rich repeat receptor-like protein kinase), Glyma.13G341400 (AGC kinase family protein), Glyma.13G331900 (60S ribosomal protein), and Glyma.13G333100 (aquaporin transporter) showed higher expression in root tissues based on publicly available transcriptome data. These results will help breeders improve soybean genetic components and enhance soybean root morphological traits using desirable alleles from wild soybeans.

Genetic control of root morphology in response to nitrogen across rapeseed diversity.

Rapeseed (Brassica napus L.) is an oil-containing crop of great economic value but with considerable nitrogen requirement. Breeding root systems that efficiently absorb nitrogen from the soil could be a driver to ensure genetic gains for more sustainable rapeseed production. The aim of this study is to identify genomic regions that regulate root morphology in response to nitrate availability. The natural variability offered by 300 inbred lines was screened at two experimental locations. Seedlings grew hydroponically with low or elevated nitrate levels. Fifteen traits related to biomass production and root morphology were measured. On average across the panel, a low nitrate level increased the root-to-shoot biomass ratio and the lateral root length. A large phenotypic variation was observed, along with important heritability values and genotypic effects, but low genotype-by-nitrogen interactions. Genome-wide association study and bulk segregant analysis were used to identify loci regulating phenotypic traits. The first approach nominated 319 SNPs that were combined into 80 QTLs. Three QTLs identified on the A07 and C07 chromosomes were stable across nitrate levels and/or experimental locations. The second approach involved genotyping two groups of individuals from an experimental F2 population created by crossing two accessions with contrasting lateral root lengths. These individuals were found in the tails of the phenotypic distribution. Co-localized QTLs found in both mapping approaches covered a chromosomal region on the A06 chromosome. The QTL regions contained some genes putatively involved in root organogenesis and represent selection targets for redesigning the root morphology of rapeseed.

Root system architecture variation among barley (Hordeum vulgare L.) accessions at seedling stage under soil acidity condition.

MAIN CONCLUSION: Soil acidity in Ethiopian highlands impacts barley production, affecting root system architecture. Study on 300 accessions showed significant trait variability, with potential for breeding enhancement. Soil acidity poses a significant challenge to crop production in the highland regions of Ethiopia, particularly impacting barley, a crucial staple crop. This acidity serves as a key stressor affecting the root system architecture (RSA) of this crop. Hence, the objective of this study was to assess the RSA traits variability under acidic soil conditions using 300 barley accessions in a greenhouse experiment. The analysis of variance indicated substantial variations among the accessions across all traits studied. The phenotypic coefficient of variation ranged from 24.4% for shoot dry weight to 11.1% for root length, while the genotypic coefficient variation varied between 18.83 and 9.2% for shoot dry weight and root length, respectively. The broad-sense heritability ranged from 36.7% for leaf area to 69.9% for root length, highlighting considerable heritability among multiple traits. The genetic advances as a percent of the mean ranged from 13.63 to 29.9%, suggesting potential for enhancement of these traits through breeding efforts. Principal component analysis and cluster analysis grouped the genotypes into two major clusters, each containing varying numbers of genotypes with contrasting traits. This diverse group presents an opportunity to access a wide range of potential parent candidates to enhance genetic variablity in breeding programs. The Pearson correlation analysis revealed significant negative associations between root angle (RA) and other RSA traits. This helps indirect selection of accessions for further improvement in soil acidity. In conclusion, this study offers valuable insights into the RSA characteristics of barley in acidic soil conditions, aiding in the development of breeding strategies to enhance crop productivity in acidic soil environments.

Response of the root anatomical structure of Carex moorcroftii to habitat drought in the Western Sichuan Plateau of China.

MAIN CONCLUSION: The anatomical structures of Carex moorcroftii roots showing stronger plasticity during drought had a lower coefficient of variation in cell size in the same habitats, while those showing weaker plasticity had a higher coefficient of variation. The complementary relationship between these factors comprises the adaptation mechanism of the C. moorcroftii root to drought. To explore the effects of habitat drought on root anatomy of hygrophytic plants, this study focused on roots of C. moorcroftii. Five sample plots were set up along a soil moisture gradient in the Western Sichuan Plateau to collect experimental materials. Paraffin sectioning was used to obtain root anatomy, and one-way ANOVA, correlation analysis, linear regression analysis, and RDA ranking were applied to analyze the relationship between root anatomy and soil water content. The results showed that the root transverse section area, thickness of epidermal cells, exodermis and Casparian strips, and area of aerenchyma were significantly and positively correlated with soil moisture content (P < 0.01). The diameter of the vascular cylinder and the number and total area of vessels were significantly and negatively correlated with the soil moisture content (P < 0.01). The plasticity of the anatomical structures was strong for the diameter and area of the vascular cylinder and thickness of the Casparian strip and epidermis, while it was weak for vessel diameter and area. In addition, there was an asymmetrical relationship between the functional adaptation of root anatomical structure in different soil moisture and the variation degree of root anatomical structure in the same soil moisture. Therefore, the roots of C. moorcroftii can shorten the water transport distance from the epidermis to the vascular cylinder, increase the area of the vascular cylinder and the number of vessels, and establish a complementary relationship between the functional adaptation of root anatomical structure in different habitats and the variation degree of root anatomical structure in the same habitat to adapt to habitat drought. This study provides a scientific basis for understanding the response of plateau wetland plants to habitat changes and their ecological adaptation strategies. More scientific experimental methods should be adopted to further study the mutual coordination mechanisms of different anatomical structures during root adaptation to habitat drought for hygrophytic plants.

Hydro-mechanical effects of vegetation on slope stability: A review.

This study explores the complex interplay between vegetation and soil stability on slopes to enhance soil-bioengineering and slope stabilization techniques. We assess the multifaceted role of vegetation in soil stabilization, examining processes such as canopy interception, stemflow, and the effects of hydrological and mechanical changes induced by root systems and above-ground plant structures. Key underlying mechanisms and their effects on stability are reported, along with the evaluation of significant plant indicators from historical research. Our review revealed that plant coverage and root architecture are critical in reducing soil erosion, with plant roots increasing soil cohesion and reducing soil detachability. Above-ground vegetation provides a protective layer that decreases the kinetic energy of raindrops and allows for higher infiltration. The importance of species-specific root traits is emphasized as pragmatic determinants of erosion prevention. Additionally, the effects of root reinforcement on shallow landslides are dissected to highlight their dualistic nature. While root-soil interactions typically increase soil shear strength and enhance slope stability, it is crucial to discriminate among vegetation types such as trees, shrubs, and grasses due to their distinct root morphology, tensile strength, root area ratio, and depth. These differences critically affect their impact on slope stability, where, for instance, robust shrub roots may fortify soil to greater depths, whereas grass roots contribute significantly to topsoil shear strength. Grasses and herbaceous plants effectively controlled surface erosion, whereas shrubs mainly controlled shallow landslides. Therefore, it is vital to conduct a study that combines shrubs with grasses or herbaceous plants. Both above-ground and below-ground plant indicators, including root and shoot indicators, were crucial for improving slope stability. To accurately evaluate the impact of plant species on slope stability reinforcement, it is necessary to study the combination of hydro-mechanical coupling with both ground plant indicators under specific conditions.

Root topological order drives variation of fine root vessel traits and hydraulic strategies in tropical trees.

Vessel traits contribute to plant water transport from roots to leaves and thereby influence how plants respond to soil water availability, but the sources of variation in fine root anatomical traits remain poorly understood. Here, we explore the variations of fine root vessel traits along topological orders within and across tropical tree species. Anatomical traits were measured along five root topological orders in 80 individual trees of 20 species from a tropical forest in southwestern China. We found large variations for most root anatomical traits across topological orders, and strong co-variations between vessel traits. Within species, theoretical specific xylem hydraulic conductivity (Kth) increased with topological order due to increased mean vessel diameter, size heterogeneity, and decreased vessel density. Across species, Kth was associated with vessel fraction in low-order roots and correlated with mean vessel diameter and vessel density in high-order roots, suggesting a shift in relative anatomical contributors to Kth from the second- to fifth-order roots. We found no clear relationship between Kth and stele: root diameter ratios. Our study shows strong variations in root vessel traits across topological orders and species, and highlights shifts in the anatomical underpinnings by varying vessel-related anatomical structures for an optimized water supply.

Deep learning-based association analysis of root image data and cucumber yield.

The root system is important for the absorption of water and nutrients by plants. Cultivating and selecting a root system architecture (RSA) with good adaptability and ultrahigh productivity have become the primary goals of agricultural improvement. Exploring the correlation between the RSA and crop yield is important for cultivating crop varieties with high-stress resistance and productivity. In this study, 277 cucumber varieties were collected for root system image analysis and yield using germination plates and greenhouse cultivation. Deep learning tools were used to train ResNet50 and U-Net models for image classification and segmentation of seedlings and to perform quality inspection and productivity prediction of cucumber seedling root system images. The results showed that U-Net can automatically extract cucumber root systems with high quality (F1_score ≥ 0.95), and the trained ResNet50 can predict cucumber yield grade through seedling root system image, with the highest F1_score reaching 0.86 using 10-day-old seedlings. The root angle had the strongest correlation with yield, and the shallow- and steep-angle frequencies had significant positive and negative correlations with yield, respectively. RSA and nutrient absorption jointly affected the production capacity of cucumber plants. The germination plate planting method and automated root system segmentation model used in this study are convenient for high-throughput phenotypic (HTP) research on root systems. Moreover, using seedling root system images to predict yield grade provides a new method for rapidly breeding high-yield RSA in crops such as cucumbers.

Relationships between root exudation and root morphological and architectural traits vary with growing season.

Plants allocate a substantial amount of C belowground for root exudates and for the construction and adjustment of root morphological and architectural traits. What relationships exist between root exudates and other root traits and these relationships change with growing season, however, remain unclear. We quantified the root exudation rate and root morphological traits, including total root length (RL), total root surface area (RS), root diameter (RD), specific root length (SRL), specific root area (SRA) and root tissue density (RTD), and architectural traits, such as branching intensity (BI), and investigated their associations during the rapidly growing season (April and August) and the slowly growing season (December) of three common native tree species, Liquidambar formosana, Michelia maudiae and Schima superba, in subtropical China. We found that the linkages of RD, SRL, SRA, RTD and BI did not change with the growing season, reflecting their highly conservative relationships. The root exudation rate varied significantly with growing season (P < 0.05) and produced various associations with other root traits at different growing seasons. During the rapidly growing season (i.e., April), the exudation rate was the highest and was positively correlated with RL. The exudation rate was the lowest during the slowly growing season (i.e., December) and was negatively associated with RL, RS and RTD. Our findings demonstrate the seasonality of the linkages of root exudation rate with other root traits, which highlights the highly plastic and complex associations of belowground root traits. These findings help to deepen our understanding of plant nutrient acquisition strategies.

The 2020 derecho revealed limited overlap between maize genes associated with root lodging and root system architecture.

Roots anchor plants in soil, and the failure of anchorage (i.e. root lodging) is a major cause of crop yield loss. Anchorage is often assumed to be driven by root system architecture (RSA). We made use of a natural experiment to measure the overlap between the genetic regulation of RSA and anchorage. After one of the most devastating derechos ever recorded in August 2020, we phenotyped root lodging in a maize (Zea mays) diversity panel consisting of 369 genotypes grown in 6 environments affected by the derecho. Genome-wide and transcriptome-wide association studies identified 118 candidate genes associated with root lodging. Thirty-four percent (40/118) of these were homologs of genes from Arabidopsis (Arabidopsis thaliana) that affect traits such as root morphology and lignin content, expected to affect root lodging. Finally, gene ontology enrichment analysis of the candidate genes and their predicted interaction partners at the transcriptional and translational levels revealed the complex regulatory networks of physiological and biochemical pathways underlying root lodging in maize. Limited overlap between genes associated with lodging resistance and RSA in this diversity panel suggests that anchorage depends in part on factors other than the gross characteristics of RSA.

Caspase 3-like protease is involved in ethylene-induced programmed cell death during aerenchyma formation in Helianthus annuus stem.

Previous research has reported that hypoxic conditions and ethylene treatments greatly trigger programmed cell death (PCD) occurrence and induce the formation of aerenchyma to adapt stress environment in Helianthus annuus stem. Caspase 3-like protease (CLP) as regulatory signals, also be involved in the process of PCD to adapt the low oxygen environment. However, the relationships between ethylene and CLP have seldom been reported. Herein, To understand the regulatory role of ethylene and CLP signaling molecules in aerenchyma formation, we investigated the effects of exogenous ethephon (ET), ethylene perception inhibitor 1-methylcyclopropene (1-MCP), and the treatment of 1-MCP + ET on morphological, physiological characteristics and aerenchyma formation in H. annuus stem. The results showed that lysigenous aerenchyma formation in H. annuus stem is induced by ET, and immunohistochemistry assay indicate CLP activity is raised at the formation stage of aerenchyma formation, and decreased at the expanding phase of aerenchyma formation. Western blotting illustrate the expression of CLP is also increased within 8 h after ethylene signaling inducing aerenchyma formation, and the activities of CLP are higher in ET treated seedlings than the control and 1-MCP treated seedlings. The same phenomenon was also observed by caspase-3 activity assay. These results revealed there is a causal and interdependent relationship between ET and CLP signaling during the process of aerenchyma formation, which regulating PCD initiation in H. annuus stem.

Influence of shrub root combinations and spacing on slope stability: study case at the Yongding River flooding regime, Langfang, China.

Research on the mechanism of plant root-soil consolidation is a current focus in research into the ecological restoration of banks. The stability of ecological banks is central to this research, and bank stability is closely related to plant combinations and spacing. Recent research on reinforced anchorage of plant roots has mainly focused on root length and angle, and on other parts of the root system, and only a few studies have examined the combination of different types of roots. In this study, a coupled slope stability assessment system is created, composed of root morphological parameters and involving calculations using the finite element model ABACUS. This paper selects the two banks of the lower reaches of the Tiantang River in the flood zone of Yongding River as the research area, and examines slope surface plants. And then the reinforcement effect of different shrub roots combinations and plant spacing are evaluated for determining the optimal shrub layout, with the aim of solving the instability problem of collapsible silty clay bank slopes and associated risks. The results indicated that when the shrub plant spacing is 0.65 m, the optimal shrub combination is Tamarix chinensis + Philadelphus incanus, and when the shrub plant spacing is 0.75 m, the optimal shrub combination is Tamarix chinensis + Euonymus alatus. The study found that the root system morphology and the fibrous roots amount at the foot of the slope can have different degrees of influence on the shallow soil stability of the silty clay slope under different shrubs plant spacing conditions.

Nitrate/ammonium-responsive microRNA-mRNA regulatory networks affect root system architecture in Populus × canescens.

BACKGROUND: Nitrate (NO3-) and ammonium (NH4+) are the primary forms of inorganic nitrogen (N) taken up by plant roots, and a lack of these N sources commonly limits plant growth. To better understand how NO3- and NH4+ differentially affect root system architecture, we analyzed the expression profiles of microRNAs and their targets in poplar roots treated with three forms of nitrogen S1 (NO3-), S2 (NH4NO3, normal), and S3 (NH4+) via RNA sequencing. RESULTS: The results revealed a total of 709 miRNAs. Among them, 57 significantly differentially expressed miRNAs and 28 differentially expressed miRNA-target pairs showed correlated expression profiles in S1 vs. S2. Thirty-six significantly differentially expressed miRNAs and 12 differentially expressed miRNA-target pairs showed correlated expression profiles in S3 vs. S2. In particular, NFYA3, a target of upregulated ptc-miR169i and ptc-miR169b, was downregulated in S1 vs. S2, while NFYA1, a target of upregulated ptc-miR169b, was downregulated in S3 vs. S2 and probably played an important role in the changes in root morphology observed when the poplar plants were treated with different N forms. Furthermore, the miRNA-target pairs ptc-miR169i/b-D6PKL2, ptc-miR393a-5p-AFB2, ptc-miR6445a-NAC14, ptc-miR172d-AP2, csi-miR396a-5p_R + 1_1ss21GA-EBP1, ath-miR396b-5p_R + 1-TPR4, and ptc-miR166a/b/c-ATHB-8 probably contributed to the changes in root morphology observed when poplar plants were treated with different N forms. CONCLUSIONS: These results demonstrate that differentially expressed miRNAs and their targets play an important role in the regulation of the poplar root system architecture by different N forms.

Maternal salinity influences anatomical parameters, pectin content, biochemical and genetic modifications of two Salicornia europaea populations under salt stress.

Salicornia europaea is among the most salt-tolerant of plants, and is widely distributed in non-tropical regions. Here, we investigated whether maternal habitats can influence different responses in physiology and anatomy depending on environmental conditions. We studied the influence of maternal habitat on S. europaea cell anatomy, pectin content, biochemical and enzymatic modifications under six different salinity treatments of a natural-high-saline habitat (~ 1000 mM) (Ciechocinek [Cie]) and an anthropogenic-lower-saline habitat (~ 550 mM) (Inowroclaw [Inw]). The Inw population showed the highest cell area and roundness of stem water storing cells at high salinity and had the maximum proline, carotenoid, protein, catalase activity within salt treatments, and a maximum high and low methyl esterified homogalacturonan content. The Cie population had the highest hydrogen peroxide and peroxidase activity along with the salinity gradient. Gene expression analysis of SeSOS1 and SeNHX1 evidenced the differences between the studied populations and suggested the important role of Na+ sequestration into the vacuoles. Our results suggest that the higher salt tolerance of Inw may be derived from a less stressed maternal salinity that provides a better adaptive plasticity of S. europaea. Thus, the influence of the maternal environment may provide physiological and anatomical modifications of local populations.

Nitrogen Uptake and Distribution in Different Chinese Cabbage Genotypes under Low Nitrogen Stress.

In order to understand the effects of low nitrogen (LN) stress on the growth and development in different genotypes of Chinese cabbage, the L40 genotype with high nitrogen utilization and the L14 genotype with LN utilization were selected as experimental materials. Field experiments and indoor hydroponic methods were used to study the different responses of two Chinese cabbage genotypes to low nitrogen levels. In this study, we also analyzed the genome-wide gene expression profiles of L40 and L14 in response to LN stress by high-throughput RNA sequencing technology. The results reveal that the L40 root system responds better to LN compared with L14. After LN stress, L40 can effectively absorb and transport NO3- and store it in the ground. It is precisely because of this characteristic of the L40 genotype that LN treatment did not have a significant effect on the chlorophyll (Chl) content and net photosynthetic rate (Pn) of the L40 Chinese cabbage compared with the L14 Chinese cabbage. These two different Chinese cabbage genotypes were shown to have differently expressed genes related to nitrate transport, auxin synthesis, and glutamate dehydrogenase synthesis. These genes function in the nitrogen pathway, which are important candidates for understanding the molecular host-response mechanisms to LN stress.

Drought-Responsive NAC Transcription Factor RcNAC72 Is Recognized by RcABF4, Interacts with RcDREB2A to Enhance Drought Tolerance in Arabidopsis.

RcNAC72, a key transcription factor that may respond to drought stress in Rosa chinensis 'Old Blush', was selected in our previous study. In the present study, we found that RcNAC72 is localized in the nucleus and is a transcriptional activator. RcNAC72 expression could be significantly induced by drought, low temperature, salt as well as abscisic acid (ABA) treatment. Analysis of the promoter revealed that multiple abiotic stress and hormone response elements were located in the promoter region. The promoter could respond to drought, low temperature, salt and ABA treatments to activate GUS gene expression. Overexpressing RcNAC72 in Arabidopsis thaliana enhanced sensitivity to ABA and tolerance to drought stress. Silencing of RcNAC72 by virus-induced gene silencing (VIGS) in rose leaves significantly reduced leaf water loss tolerance and leaf extension capacity. Physical interaction of RcNAC72 with RcDREB2A was shown by means of the yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) assays. RcABF4 was demonstrated to be able to bind to the promoter of RcNAC72 by means of the yeast one-hybrid (Y1H) assay. These results provide new insights into the regulatory network of RcNAC72 response to drought stress in roses.

Using anatomical traits to understand root functions across root orders of herbaceous species in a temperate steppe.

Root anatomical traits play crucial roles in understanding root functions and root form-function linkages. However, the root anatomy and form-function linkages of monocotyledonous and dicotyledonous herbs remain largely unknown. We measured order-based anatomical traits and mycorrhizal colonization rates of 32 perennial herbs of monocotyledons and dicotyledons in a temperate steppe. For monocots, relative constant proportion of cortex and mycorrhizal colonization rates, but increased cell-wall thickening of the endodermis and proportion of stele were observed across root orders, indicating a slight reduction in absorption capacity and improvement in transportation capacity across orders. For dicots, the cortex and mycorrhizal colonization disappeared in the fourth-order and/or fifth-order roots, whereas the secondary vascular tissue increased markedly, suggesting significant transition of root functions from absorption to transportation across root orders. The allometric relationships between stele and cortex differed across root orders and plant groups, suggesting different strategies to coordinate the absorption and transportation functions among plant groups. In summary, our results revealed different functional transition patterns across root orders and distinct strategies for coordinating the absorption and transportation of root system between monocots and dicots. These findings will contribute to our understanding of the root form and functions in herbaceous species.

GmWRKY46, a WRKY transcription factor, negatively regulates phosphorus tolerance primarily through modifying root morphology in soybean.

Phosphorus (P) deficiency affects soybean growth and development, resulting in significant reduction of yields. However, the regulatory mechanism of P deficiency tolerance in soybean is still largely unclear. WRKY transcription factors are a family of regulators involved in a variety of abiotic stresses in plants while rarely reported in P deficiency. Here, we demonstrated that a soybean GmWRKY46 gene, belonging to group III of WRKY TF family, was involved in the regulation of P deficiency tolerance in soybean. The expression of GmWRKY46 in low P sensitive soybean varieties was significantly higher than that in tolerant soybean varieties. It was primarily expressed in roots and strongly induced by P deprivation. GmWRKY46 was localized in the nucleus. Compared with the control expressing the empty vector, overexpression of GmWRKY46 in soybean hairy roots exhibited more sensitive phenotypes to low P stress, while the RNA interfered GmWRKY46 significantly enhanced P deficiency tolerance by increasing the proliferation, elongation and P absorption efficiency of hairy roots. Expression patterns of a number of P-responsive genes (GmPht1;1, GmPht1;4, GmPTF1, GmACP1, GmPAP21 and GmExpansin-A7) were altered in both overexpression and gene silenced plants. The results provided a novel insight into how soybean responds to low P stress and new gene that may be used to improve soybean low P tolerance through gene editing approach.

Optical Clearing of Plant Tissues for Fluorescence Imaging.

It is challenging to directly observe the internal structure of multi-layered and opaque plant specimens, without dissection, under a microscope. In addition, autofluorescence attributed to chlorophyll hampers the observation of fluorescent proteins in plants. For a long time, various clearing reagents have been used to make plants transparent. However, conventional clearing reagents diminish fluorescent signals; therefore, it has not been possible to observe the cellular and intracellular structures with fluorescent proteins. Reagents were developed that can clear plant tissues by removing chlorophyll while maintaining fluorescent protein stability. A detailed protocol is provided here for the optical clearing of plant tissues using clearing reagents, ClearSee (CS) or ClearSeeAlpha (CSA). The preparation of cleared plant tissues involves three steps: fixation, washing, and clearing. Fixation is a crucial step in maintaining the cellular structures and intracellular stability of fluorescent proteins. The incubation time for clearing depends on the tissue type and species. In Arabidopsis thaliana, the time required for clearing with CS was 4 days for leaves and roots, 7 days for seedlings, and 1 month for pistils. CS also required a relatively short time of 4 days to make the gametophytic leaves of Physcomitrium patens transparent. In contrast, pistils in tobacco and torenia produced brown pigment due to oxidation during CS treatment. CSA reduced the brown pigment by preventing oxidation and could make tobacco and torenia pistils transparent, although it took a relatively long time (1 or 2 months). CS and CSA were also compatible with staining using chemical dyes, such as DAPI (4',6-diamidino-2-phenylindole) and Hoechst 33342 for DNA and Calcofluor White, SR2200, and Direct Red 23 for the cell wall. This method can be useful for whole-plant imaging to reveal intact morphology, developmental processes, plant-microbe interactions, and nematode infections.

UCP1 and AOX1a contribute to regulation of carbon and nitrogen metabolism and yield in Arabidopsis under low nitrogen stress.

Nitrogen (N) availability is a critical factor for plant development and crop yield, and it closely correlates to carbon (C) metabolism. Uncoupling protein (UCP) and alternative oxidase (AOX) exhibit a strong correlation with N and C metabolism. Here, we investigated the functions of UCP1 and AOX1a using their mutants and complementation lines in Arabidopsis adaptation to low N. Low N markedly increased AOX1a and UCP1 expression, alternative pathway capacity and UCP activity. Eight-day-old aox1a/ucp1 seedlings were more sensitive to low N than Col-0 and single mutants, exhibiting lower primary root length and higher anthocyanin accumulation. The net photosynthetic rate, electron transport rate, PSII actual photochemical efficiency, stomatal conductance and carboxylation efficiency were markedly decreased in ucp1 and aox1a/ucp1 compared to those in Col-0 and aox1a under low N stress; comparatively, chlorophyll content and non-photochemical quenching coefficient were the lowest and highest in aox1a/ucp1, respectively. Nitrate acquisition rate was accelerated in aox1a/ucp1, but its transport activity was decreased, which resulted in low nitrate content and nitrate reductase activity under low N condition. The C/N ratio in seeds, but not in leaves, is higher in aox1a/ucp1 than that in Col-0, aox1a and ucp1 under low N condition. RNA-seq analysis revealed that many genes involved in photosynthesis and C/N metabolism were markedly down-regulated in aox1a/ucp1 under low N stress. These results highlight the key roles of UCP1 and AOX1a in modulating photosynthetic capacity, C/N assimilation and distribution under low N stress.

Potassium transporter TRH1/KUP4 contributes to distinct auxin-mediated root system architecture responses.

In plants, auxin transport and development are tightly coupled, just as hormone and growth responses are intimately linked in multicellular systems. Here we provide insights into uncoupling this tight control by specifically targeting the expression of TINY ROOT HAIR 1 (TRH1), a member of plant high-affinity potassium (K+)/K+ uptake/K+ transporter (HAK/KUP/KT) transporters that facilitate K+ uptake by co-transporting protons, in Arabidopsis root cell files. Use of this system pinpointed specific root developmental responses to acropetal versus basipetal auxin transport. Loss of TRH1 function shows TRHs and defective root gravitropism, associated with auxin imbalance in the root apex. Cell file-specific expression of TRH1 in the central cylinder rescued trh1 root agravitropism, whereas positional TRH1 expression in peripheral cell layers, including epidermis and cortex, restored trh1 defects. Applying a system-level approach, the role of RAP2.11 and ROOT HAIR DEFECTIVE-LIKE 5 transcription factors (TFs) in root hair development was verified. Furthermore, ERF53 and WRKY51 TFs were overrepresented upon restoration of root gravitropism supporting involvement in gravitropic control. Auxin has a central role in shaping root system architecture by regulating multiple developmental processes. We reveal that TRH1 jointly modulates intracellular ionic gradients and cell-to-cell polar auxin transport to drive root epidermal cell differentiation and gravitropic response. Our results indicate the developmental importance of HAK/KUP/KT proton-coupled K+ transporters.